Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant vascular disorder, characterized by spontaneous recurrent nosebleeds, mucocutaneous telangiectases, and arterio- venous malformations (AVMs) in the brain, lung, liver or GI tract. While reduced expression of either Endoglin (ENG) or Activin receptor-like kinase 1 (ALK1) has shown to be associated HHT, the precise pathogenetic mechanisms underlying HHT remain elusive; and thus, while management options for HHT patients are well established, treatment options for this malady is currently lacking. The ultimate goal of this project is to develop novel therapeutic reagents for treating HHT. To reach this goal we set out the following five stepwise goals: 1) Development of mouse models that reproduce clinical features of vascular malformations found in HHT patients; 2) Elucidation of pathogenetic mechanisms that underlie the vascular malformations using the animal model; 3) Discovery of potential therapeutic target that can prevent or reverse the vascular malformations based on the mechanism; 4) Preclinical validation of effects of the potential therapies using the animal models; 5) Clinical trials of validated therapies through multi-HHT centers of excellence. In the past funding period, we have focused on the first goal: development of reliable animal models for HHT study. As shown in the progress report and preliminary data sections below, we have accomplished this goal. In the next funding period, we will focus on goals 2-4. We hope that by the end of next funding period, we would be able to bring solid therapeutic options to carry out clinical trials. We will utilize the advanced genetic models in combinations with a state-of-the-art intravital vascular imaging technology, in vitro assays, and systems biological approaches for preclinical studies and for discovering novel therapeutic targets for preventing AVM development. To reach this goal we propose the following three specific aims. Aim 1 is to investigate cellular events during AVM formation at a subcellular level. Deep-penetrating and noninvasive multiphoton fluorescence microscopy will be employed to capture 3-dimensional time lapse images blood vessels at subcellular level during AVM formation. Combination of multiphoton microscopy with the current hyperspectral imaging system will greatly improve our animal model to facilitate our understanding of the progression of AVM formations. Aim 2 is to establish preclinical animal models for evaluating potential drugs targeting AVM pathogenesis. Three therapeutic drugs that have been used for treating HHT patients (tranexamic acid, thalidomide, and a VEGF-trap) will be evaluated using our animal models. Aim 3 is to determine molecular pathways and downstream effector genes of ALK1 signaling pertinent to angiogenesis and AVM formation. We will establish pulmonary ECs from tamoxifen-inducible Alk1-conditional knockout mice. Using these EC lines, we will investigate cellular, biochemical and molecular characteristics of Alk1-deficient ECs. We will also perform microarray analyses using RNAs isolated from tissues of multiple HHT animal models and mutant cells to determine downstream target genes of ALK1 signaling that are relevant for HHT pathogenesis. The results from the proposed experiments will greatly facilitate development of novel therapeutic targets for HHT, especially for controlling nosebleeds and GI bleedings, and benefit HHT patients. The proposed studies will also accelerate the advances in general knowledge of AVM biology. In addition, these studies may uncover a novel control mechanism of angiogenesis, which will be applicable for a variety of pathological conditions and diseases, including retinopathy, wound healing, nephropathy, and cancer. In a basic science aspect proposed studies will enhance general knowledge about endothelial cell biology and TGF-2 signaling. PUBLIC HEALTH RELEVANCE: People with hereditary hemorrhagic telangiectasia (HHT), an inherited rare blood disease, contains arteriovenous malformation (AVM)-arteries and vein are directly connected without capillaries in between them-in various organs in a very high frequency compared with normal individuals. The overall goal of the proposal is to find out 'why AVMs develop in HHT patients' and 'how we can inhibit formation (and growth) of AVMs' using mouse models. Results from this study map provide important novel insights for developing therapeutic reagents for controlling development of AVMs.